30 million years of biofilm preservation in Rio Tinto's acidic iron-sulfate system

The Río Tinto system (SW Spain) provides a unique natural archive for evaluating long-term preservation of complex microbial biofilms under extreme acidic and oxidizing conditions. Here we demonstrate that mineralized biofilms preserved over ~30 million years retain not only molecular biosignatures, but also the ecological diversity characteristic of modern Río Tinto biofilm communities. Using high-resolution Time-of-Flight Secondary Ion Mass Spectrometry (ToF-SIMS), we analyze iron–sulfate deposits from the Upper Gossan (~30 Ma), Upper Terrace (~2.1 Ma), and Intermediate Terrace (~7.5 ka). Upper Gossan deposits preserve spatially coherent fungal- and algal-derived lipid assemblages, including ceramides, diacylglycerides, wax esters, polyunsaturated fatty acids, and sulfated sterols, directly mirroring biofilm architectures and taxonomic compositions observed in the modern river. Terrace deposits record progressively reduced molecular fidelity linked to seasonal hydrology, while Intermediate Terrace materials show near-complete lipid loss. These results demonstrate that under hyperacidic conditions, iron–sulfate mineralization can preserve biofilm diversity and community structure over geological timescales, providing a powerful analog for biosignature preservation on early Earth and Mars.